Bone as an endocrine organ: the biological role and mechanisms of action of osteocalcin in the vasculature

这个迷你项目将解决两个目标：(1)骨钙素对人肠系膜动脉的直接血管作用；(2)骨钙素对内皮细胞的影响。(1)在人的肠系膜动脉中，我们将使用丝状肌图术(O‘Sullivan等人，2004；2005；2006；2009)，检查骨钙素在应用于基线音调时，是导致预先收缩的动脉的血管松弛，还是导致动脉的血管收缩。我们还将通过将动脉暴露于骨钙素，药物预收缩，然后对内皮依赖性和非依赖性血管松弛药进行浓度反应曲线，从生理学上检验与生理相关的骨钙素浓度对后续血管扩张功能的影响。骨钙素对去甲肾上腺素、血管紧张素II和内皮素-1的血管收缩反应的影响也将被检测。(2)到目前为止，还没有研究过骨钙素对内皮细胞的影响，尽管最近的一项研究表明，骨钙素通过缺氧诱导因子1信号，刺激血管平滑肌细胞中骨软骨性转录因子的表达(Idelevich等人，2011年)。学生将研究骨钙素对人内皮细胞系的影响，以确定骨钙素是否影响内皮细胞功能的一系列特征，包括内皮细胞增殖(四甲基偶氮唑盐比色法)、细胞毒性(乳酸脱氢酶试验)、血管黏附分子、血管细胞黏附分子和钙化标记物(碱性磷酸酶含量或钙染色)。

The mini project will address two aims; (1) what are the direct vascular effects of osteocalcin in human mesenteric arteries, and (2) what are the effects of osteocalcin on endothelial cells. (1) Using wire myography (O'Sullivan et al., 2004; 2005; 2006; 2009) in human mesenteric arteries we will examine whether osteocalcin causes vasorelaxation of pre-constricted arteries or causes vasoconstriction of arteries when applied to baseline tone. We will also examine physiologically the effect of physiologically-relevant concentrations of osteocalcin on subsequent vasodilator function by exposing arteries to osteocalcin, pre-constricting pharmacologically and then performing concentration responses curves to endothelium-dependent and independent vasorelaxants. The effect of osteocalcin on vasoconstrictor responses to noradrenaline, angiotensin II and endothelin-1 will also be examined. (2) No studies to date have examined the effects of osteocalcin on endothelial cells, although a recent study suggested that osteocalcin, via hypoxia-inducible factor 1 signalling, stimulates expression of osteochondrogenic transcription factors in VSMCs (Idelevich et al., 2011). Students will investigate the effects of osteocalcin on a human endothelial cell line to establish whether osteocalcin affects a number of features of endothelial cell function, including endothelial cell proliferation (MTT assay), cytotoxicity (LDH assay), the expression of vascular adhesion molecule, vCAM (by ELISA), and markers of calcification (ALP content or calcium staining).